The laboratory uses a broad range of molecular, biochemical and biophysical research tools centered around X-ray crystal structure determination to understand the mechanism of chromatin recognition and assembly and post-translational histone and protein modification in the regulation of gene expression; and kinase signaling pathways. The laboratory is particularly interested in gene regulatory proteins and their upstream signaling kinases that are aberrantly regulated in cancer and age-related metabolic disorders such as type II diabetes and obesity, and the use of high-throughput small molecule screening and structure-based design strategies towards the development of protein-specific small-molecule probes to be used to further interrogate protein function and for development into therapeutic agents.

Chromatin recognition and assembly and histone modification in gene regulation. DNA within the eukaryotic nucleus is compacted into chromatin containing histone proteins and its appropriate regulation orchestrates all DNA-templated reactions such as DNA transcription, replication, repair, mitosis, and apoptosis. Among the many proteins that regulate chromatin, the proteins that recognize DNA, assemble chromatin, called histone chaperones, and that modify the histones through the addition or removal of functional groups such as acetyl, methyl or phosphate play important roles. We are studying the DNA binding proteins p53, FoxO and the Gal4 family; the histone chaperones HIRA, Asf1, Vps75 and their associated factors; and the family of histone acetyltransferase (HAT) and histone deacetylase (HDAC) enzymes. We are particularly interested in how DNA binding proteins navigate the recognition of their cognate DNA targets, how histone chaperones coordinate the assembly of distinct chromatin complexes correlated with different DNA regulatory processes, and how histone modification enzymes link catalysis to their substrate specific activities for their respective biological activities. More recently, we have been studying how the binding of accessory and regulatory protein subunits regulates the various activities of these proteins and in some cases we are developing small molecule protein specific inhibitors.

Enzymes associated with aging and age-related disorders.Sirtuin enzymes are NAD+-dependent histone and protein deactylases and/or ADP-ribotransferases that have been implicated in the regulation of gene expression, cellular aging, adipogenesis, type II diabetes and several neurodegenerative disorders. We have determined the structure of these enzymes in several liganded forms and have developed novel small molecule sirtuin inhibitors. Together with associated biochemical studies, these studies have provided insights into the mode of catalysis and substrate-specific recognition by this protein family and have illuminated new avenues for small molecule effector design. We are currently working towards understanding the factors that distinguish different sirtuin proteins and how the functions of these proteins are modulated by other protein factors. We are also pursuing structure/function studies of other proteins that are implicated in aging and age-related disorders.

Tumor suppressors and oncoproteins. We are carrying out biochemical and structural studies on the tumor suppressor proteins pRb, p53 and p300/CBP, both alone and in complex with their relevant protein targets. We are also interested in the mode of inactivation of these tumor suppressors by the viral oncoproteins E7 and E6 from human papillomavirus (HPV), the etiological agent for cervical cancer, and Adenovirus (Ad) E1A. We are also combining structural studies with small molecule screening to prepare small molecule HPV-E7 and for HPV-E6 inhibitors. Most recently we have begun to exploit structure-based design strategies to develop inhibitors of oncogenic kinases, such as PI3K, BRAF and PAK1 implicated in melanoma and other cancers. Our goal for these studies is to derive functional and structural information that will lead to the design of small molecule compounds that may have therapeutic applications.

Tumor suppressors and viral oncoproteins- We are pursuing biochemical and structural studies on the tumor suppressor proteins p18INK4c, pRb, p53 and p300/CBP, both alone and in complex with their relevant protein targets. The activity of pRb is inhibited by several known DNA viral oncoproteins, including human papillomavirus (HPV) E7, the etiological agent for cervical cancer, and Adenovirus (Ad) E1A. We have most recently characterized the binding properties of pRb to HPV-E7 and Ad-E1a and are now determining their structures both alone and in complex with pRb. Our goal for these studies is to derive functional and structural information that will lead to the design of small molecule compounds that may have clinical applications against cancer.

Protein-DNA recognition- As a model to understand sequence-specific DNA recognition by transcriptional regulatory proteins, we are studying the structure and function of three families of DNA binding proteins, the fungal specific Zn2Cys6 binuclear cluster proteins, the higher eukaryotic Ets proteins and p53. We have determined several structures of these proteins either alone or in complex with their associated DNA targets and are continuing to use these proteins as a model to understand DNA recognition by protein and protein complexes. With regard to p53, we are studying its unique mode of DNA recognition and are developing structure-based strategies for the repair of tumor-derived p53 mutations.

Since the lac operon has been the paradigm for gene regulatory systems, our efforts have been focused on obtaining structural information on the repressor operator complex. To this end, a series of DNA binding domain fragments and variants have been cloned and expressed for solution multinuclear multidimensional NMR analysis, both as proteins and protein DNA complexes. In collaboration with M. Lewis of the Department of Biochemistry and Biophysics, the three-dimensional structures of (1) lac repressor alone, (2) lac repressor with inducer, and (3) complexes of intact tetrametic lac repressor, and operator DNA have been solved.

An interesting result from our group is the solution structure of A-tract DNA. This variation from the average B form has been studied for two decades. Several X-ray structures by other groups have been not consistent with the body of solution properties of this family of DNA sequences. Our NMR solution structure shows that the bend In the helix axis is actually 90 degrees away from the bend plane in the crystal structures. The variation of nucleic acid structures as a function of sequence and solvent conditions is an important consideration. In related experiments, we have exploited the use of fluorescence depolarization of bound ethidium bromide to investigate hydrodynamic size and shape of RNA structures, e.g., tetraloops and ribozymes. These issues have also become important as we are finding variations in mRNA accessibility to antisense probes.

In collaboration with the late Alan Gewirtz we demonstrated that anti-sense oligonucleotides actually find their complementary sequences in the cell.

Ph.D., Cornell University (1989) B.A., University of California, Berkeley (1984)

Research Interests:

In my research group, we use a combination of analytical theory and numerical simulation to study problems in soft matter physics ranging from jamming in glassforming liquids, foams and granular materials, to biophysical self-assembly in actin structures and other systems. The idea of jamming is that slow relaxations in many different systems, ranging from glassforming liquids to foams and granular materials, can be viewed in a common framework. For example, one can define jamming to occur when a system develops a yield stress or extremely long stress relaxation time in a disordered state. According to this definition, many systems jam. Colloidal suspensions of particles are fluid but jam when the pressure or density is raised. Foams and emulsions (concentrated suspensions of deformable bubbles or droplets) flow when a large shear stress is applied, but jam when the shear stress is lowered below the yield stress. Even molecular liquids jam as temperature is lowered or density is increased this is the glass transition. We have been testing the speculation that jamming has a common origin in these different systems, independent of the control parameter varied. On the biophysical side, our research has been motivated recently by the phenomenon of cell crawling. When a cell crawls, its cytoskeleton--a network of filaments (primarily composed of the protein actin) that maintains the mechanical rigidity of the cell and gives the cell its shape--must reorganize in structure. This reorganization is accomplished via polymerization, depolymerization and branching of actin filaments, as well as by crosslinking the filaments together with "linker" proteins. The morphology of the resulting structure is of special interest because it determines the mechanical properties of the network. We are developing dynamical descriptions that capture morphology. In addition, we are exploring models for how actin polymerization gives rise to force generation at the leading edge.

Criegee intermediates: Research in the Lester laboratory is currently focused on the photo-induced chemistry of Criegee intermediates. Alkene ozonolysis is a primary oxidation pathway for alkenes emitted into the troposphere and an important mechanism for generation of atmospheric OH radicals, particularly in low light conditions, urban environments, and heavily forested areas. Alkene ozonolysis proceeds through Criegee intermediates, R1R2COO, which eluded detection until very recently. In the laboratory, the simplest Criegee intermediate, CH2OO, and methyl-substituted Criegee intermediates, CH3CHOO and (CH3)2COO, have now been generated by an alternative synthetic route and detected by VUV photoionization. This laboratory has further shown that UV excitation of the Criegee intermediates on a strong π*←π transition induces photochemistry, which involves multiple coupled excited state potentials and yields both O3P and O1D products. This group has also demonstrated that IR excitation of methyl-substituted Criegee intermediates in the CH stretch overtone region initiates unimolecular decay. The latter enables direct examination of the hydrogen transfer reaction leading to OH products, which is a key non-photolytic source of OH radicals in the atmosphere.

Hydrogen trioxide radical: This laboratory obtained the first infrared spectrum of the hydrogen trioxide (HOOO) radical, an intermediate invoked in the H + O3 and O + HO2 atmospheric reactions as well as the efficient vibrational relaxation of OH radicals by O2. There had been much debate in the literature as to whether HOOO is stable or metastable with respect to the OH + O2 limit, as well as the relative stability of the cis and trans conformers. We have characterized the geometric structure, vibrational frequencies, and stability of the cis and trans conformers of HOOO and its deuterated analog. In particular, by measuring the OH product state distribution following IR excitation of HOOO, we have directly determined the stability of trans-HOOO and shown that is much greater than prior estimates. As a result, HOOO may act as temporary sink for OH radicals and be present in measurable concentrations in the Earth's atmosphere. The experimental stability indicates that 25% of the OH radicals in the vicinity of the tropopause may be bound to O2, rather than free OH radicals. Studies of combination bands in the fundamental OH stretch region reveal nearly all other vibrational modes of trans- and cis-HOOO. We have subsequently derived a torsional potential from our spectroscopic data to obtain the relative stability of the cis and trans conformers and isomerization barrier, which are critical for atmospheric modeling of HOOO.

IR action spectrum of cis- and trans-HOOO in the OH overtone region (left), and fraction of atmospheric OH predicted to exist as HOOO (right).

Dynamical signatures of quenching: Collisional quenching of electronically excited OH A2Σ+ radicals has been extensively investigated because of its impact on OH concentration measurements in atmospheric and combustion environments. Yet little is known about the outcome of these events, except that they facilitate the efficient removal of OH population from the excited A2Σ+ electronic state by introducing nonradiative decay pathways. The quenching of OH A2Σ+ by H2 and D2 has emerged as a benchmark system for studying the nonadiabatic processes that lead to quenching. Theoretical calculations indicate that a conical intersection funnels population from the excited to ground electronic surfaces. Our studies examined the Doppler profiles for the H/D-atom products of reactive quenching, which show that most of the excess energy results in vibrational excitation of ‘hot’ water products. Our work also focused on characterizing the nonreactive quenching process, where OH X2Π products are generated with a remarkably high degree of rotational excitation and lambda-doublet specificity. The OH quantum state distribution directly reflects the anisotropy and A′ symmetry of the conical intersection region. We also demonstrated for H2 and D2 collision partners that reaction accounts for nearly 90% of the quenched products. These distinctive dynamical signatures of passage through a conical intersection region have sparked intense theoretical interest in this system.

We gratefully acknowledge financial support from the National Science Foundation under Grant No. NSF CHE-1362835 and the Chemical Sciences, Geosciences and Biosciences Division, Office of Basic Energy Sciences, Office of Science, U.S. Department of Energy under Grant No. DE-FG02-87ER13792.

The central theme of research in my laboratory is the rational design of new methods and catalysts for use in organic synthesis. As well as using traditional screening and development approaches, we employ several novel computational tools for the discovery and optimization of new reagents and catalysts. These new synthetic methods comprise the key steps in our total synthesis strategies to a variety of important pharmaceutical agents and natural products.

Asymmetric Oxidative C-C Bond Forming Reactions: The development of chiral catalysts for oxidative C-C bond formation is a major focus in our laboratory. In addition to the substantial potential for developing biomimetic synthetic approaches to a variety of natural products, such transformations are appealing in that C-H bonds are directly transformed to C-C bonds with an inexpensive oxidant, molecular oxygen.

To this end, we developed 1,5-diaza-cis-decalin copper complexes, the catalysts of choice for the oxidative asymmetric biaryl coupling of 2-naphthol derivatives. Study of the mechanism has allowed the development of new reaction methods as well as couplings of highly functionalized 2-naphthols. With this ability, we have completed the first asymmetric synthesis of the natural product nigerone. The first total syntheses of the complex natural products cercosporin and hypocrellin have also been accomplished. These structurally novel compounds display promising photodynamic therapy profiles in cancer treatment. Future goals include exploiting the oxidative biaryl coupling method in the synthesis of chiral bisanthraquinone and naphthodianthrone natural products.

Reactions of α-Ketoesters and Derivatives: We have described bifunctional salen-derived catalysts that contain electronically decoupled Lewis acid and Lewis base sites. This electronic decoupling permits generation of optimally active catalysts as both the Lewis acid and Lewis base can be maximized without quenching each other. These catalysts are particularly effective for the very difficult asymmetric alkylation of α-ketoesters and α-iminoesters to yield α-hydroxy and α-amino acid adducts in enantiomerically pure form. Further studies with α-iminoesters have revealed an umpolung addition pathway allowing addition of nucleophiles to imine nitrogens. We have exploited this reactivity pattern to develop a three-component coupling that generates highly functionalized α-amino acid derivatives.

Computer-Aided Design of Chiral Auxiliaries and Catalysts:Diastereo- and enantioselective chemical reactions are essential components for the efficient synthesis of complex chiral targets. We have generated several computational tools to assist researchers in designing and optimizing chiral catalysts including database searching and functionality mapping. In addition, we have developed semi-empirical quantum mechanical quantitative structure selectivity (QSSR) relationships for accurate and precise enantiomeric excess predictions of chiral catalysts. In one example, we correlated the structures of various beta-amino alcohol catalysts to their enantioselectivities in the asymmetric addition of diethylzinc to benzaldehyde. With our method the selectivities of new catalysts were also calculated. Subsequent chemical synthesis and analysis of the new catalysts indicated that the model was very useful and easily distinguished catalysts of low, moderate, and high selectivity.

B.S.E. Materials Science and Engineering, University of Pennsylvania 1991

B.A. Mathematics, University of Pennsylvania 1991

Research Interests:

Cherie earned both a B.S.E. in Materials Science and Engineering and a B.A. in Mathematics from the University of Pennsylvania in 1991. In 1996, she received her Ph.D. in Electronic Materials from MIT. Her thesis work focused on the self-assembly of close packed solids of semiconductor nanocrystals and the unique electronic and optical properties that arise from cooperative interactions between neighboring nanocrystals. In 1996, Cherie went to Bell Laboratories as a Postdoctoral Fellow where she built a scanning confocal Raman microscope to study the mechanistics of hologram formation in multicomponent photopolymers. In 1998 she joined IBM's T. J. Watson Research Center where she most recently managed the "Molecular Assemblies and Devices Group." In January, 2007 Cherie joined the faculty of the University of Pennsylvania's Departments of Electrical and Systems Engineering and Materials Science and Engineering as an associate professor. In addition she assumed the position as the Director of the University's Nanofabrication facility.

Cherie was selected by the American Chemical Society Women Chemist Committee in 2002 as one of 12 "Outstanding Young Woman Scientists who is expected to make a substantial impact in chemistry during this century." She was featured by the American Physical Society in "Physics in Your Future" and in 2000 chosen by the MIT Technology Review TR10. In 2005, she received IBM's Outstanding Technical Achievement award. She is on the editorial board of American Chemical Society's journal "Nano Letters" and serves on the Materials Research Society's Board of Directors and the NSF advisory board for the US Summer School in Condensed Matter and Materials Physics.

Chemical and physical properties of molecular, supramolecular, and nanoscale assemblies and devices; intramolecular, intermolecular, and interfacial charge and excitonic transport and interactions for the application of molecular and nanoscale materials in transistors and memory devices, photovoltaic devices, and chemical and biological sensors.

Investigations carried out in our laboratory encompass a wide range of interests in synthetic organic chemistry including heterocyclic and medicinal chemistry.

Current efforts are in the following areas: (1) synthesis and chemistry of five-membered heterocycles and natural products containing such units; (2) synthesis and chemistry of fungal metabolites; (3) synthesis and chemistry of cyclopeptide alkaloids; (4) synthesis of biologically important depsipeptides; (5) synthesis of novel ninhydrins; (6) synthesis of anti- angiogenic agents.

Utilization of D-ribonolactone and other sugars as precursors in the synthesis of several structurally challenging molecules is currently underway in our laboratory.

The synthesis of naturally occurring fungal metabolites containing a common hexasubstituted aromatic ring but different side chains such as colletochlorin D, ascofuranone and ascochlorin are another area of interest. The biological activities of those natural products range from high hypolipidemic action to anticancer and antiprotozoan activity.

Cyclopeptide alkaloids are natural products found in many plant families. A broad program aimed at developing methodology for the synthesis of the most commonly found thirteen- and fourteen-membered ring cyclopeptide alkaloids is currently underway. Sanjoin, used in Chinese folk medicine is one of our targets. Other antitumor cyclic peptides provenient from plants, the astins, are also under investigation.

Didemnins are a new class of depsipeptides isolated from a Carribean tunicate of the family Didemnidae, a species of the genus Trididemnum. These cyclic peptides have shown highly active antiviral and antitumor agents. The synthetic studies carried out in our laboratory have produced synthetic and spectral evidence for the absolute configuration of the asymmetric centers of the hydroxyisovalerylpropionyl (HIP) unit of the macrocycle, thereby requiring a revision of the original stereochemistry. The stereocontrolled total synthesis of these natural products has already been accomplished. The synthesis of several beta-turn mimics and constrained analogs are under investigation. Because of a broad program to develop efficient synthetic routes to the didemnins, other cyclodepsipeptides have been chosen as the next targets. The choice of these compounds was not only based on their relationship to didemnins but also on previous synthetic studies of products originating from polyketide biosynthesis, and earlier investigations of carbohydrates.

Novel ninhydrins are being synthesized as reagents for the detection of amino acids.

We have found that sulfated beta-cyclodextrin mimicked heparin advantageously. This effective synthetic product is of utmost importance in the control of angiogenesis and has other important applications in medicine. This recent discovery uncovers a new class of anti-angiogenic agents, consisting of a hydrophilic carrier and a hydrophobic angiostat, and offers a unique opportunity for the development of chemical technologies which will have important applications in the bio- and medical sciences. We are therefore continuing these studies with several goals in mind. We are investigating new and more effective carriers, we are designing single species that contain both the angiostat and carrier, and we are looking for new and more effective angiostats.

The focus of our research is to study how proteins fold from random or quasi-random coils to their biologically functional conformations. We are particularly interested in the kinetic aspects of the folding mechanisms. Novel laser spectroscopic methods are being used and developed to study the early folding events and folding intermediates.

Fast events in protein folding

Understanding how folding proceeds at early time is apparently essential to the elucidation of the entire folding mechanism. To access and characterize the early folding events requires a fast initiation method and a probe that has structural specificity. Our general approach is to use novel laser-induced temperature-jump and fast-mixing techniques to initiate refolding/unfolding on nanosecond or microsecond timescales, and use time-resolved infrared and fluorescence spectroscopies to probe the subsequent folding dynamics and structural ordering along the folding/unfolding pathways. This approach provides not only fast time resolution, but also the necessary structural sensitivity, since both infrared and fluorescence are well-established conformation probes. Recent works involve the study of the helix-coil transition, helix-helix interaction, and ß-sheet formation.

Single-molecule study of protein conformation dynamics

Recently, a new view of the kinetics of protein folding has emerged based on the new conceptual framework of statistical mechanical models, replacing the pathway concept with the broader notion of rugged energy landscapes. The heterogeneity in folding kinetics therefore can be realized as a result of the motions of an ensemble of protein conformations on the rugged energy hypersurface that is biased towards the native state, analogous to parallel diffusion-like processes. Studying folding dynamics statistically using single-molecule techniques will provide unique information regarding a protein's folding energy landscape, which may not be obtained by conventional ensemble studies since the conventional measurements of molecular dynamics in the condensed phase represent only averages over large numbers of molecules and events. Currently, confocal fluorescence spectroscopy and microscopy are being used to study protein spontaneous fluctuation and folding dynamics at single-molecule level.

Selected Publications:

S. Mukherjee, P. Chowdhury and F. Gai, “Infrared study of the effect of hydration on the amide I band and aggregation properties of helical peptides,” J. Phys. Chem. B 2007, 111, 4596.

From a knowledge of the interactions among molecules, it is possible in principle to predict the structure and the thermodynamic properties of materials as well as the dynamics of molecular processes. The overall objective of our research program is twofold: to evaluate the potential energies of intermolecular interactions for various systems as accurately as possible and to study by means of statistical mechanics the influence of these potentials of intermolecular force on the structure and properties of macroscopic systems.